Numerous studies have demonstrated myocyte apoptosis during myocardial infarction, ischemia-reperfusion injury, and chronic heart failure. Despite these observations, the two most critical questions in the field remain unexplored: 1) What is the precise molecular mechanism of apoptosis in cardiac myocytes? 2) To what extent does myocyte apoptosis contribute to myocardial dysfunction in these disease states? The research program described herein addresses both of these interrelated questions. To facilitate a molecular genetic analysis, models of myocardial infarction and ischemia-reperfusion injury have been developed and characterized in the mouse. Using genetically altered mice, we have tested the necessity of proteins that mediate apoptosis in non-cardiac contexts for apoptosis during myocardial infarction. These studies have shown that one such protein, p53, while present in ischemic cardiac myocytes and sufficient to induce apoptosis in these cells, is not required for myocyte apoptosis. This result suggests that the apoptotic program in complex pathophysiologic states can be activated by multiple, redundant signaling pathways. In contrast, the caspases, a family of cysteine proteases, are components of the final common pathway for apoptosis in all metazoan cells from worm to mammal. Indeed we have shown that pharmacologic blockade of these enzymes markedly inhibits myocyte apoptosis during myocardial infarction in vivo. The potential significance of this result is two-fold: First, caspase inhibition may provide a direct means to determine the contribution of myocyte apoptosis to myocardial dysfunction. Second, caspase inhibition may provide a new therapeutic approach to ischemic heart disease and heart failure. We now propose to deepen our understanding of the mechanism and significance of cardiac myocyte apoptosis through the following specific aims: 1. To determine which caspases are expressed in adult cardiac myocytes and undergo proteolytic activation during myocardial infarction and ischemia-reperfusion injury. 2. To block myocyte apoptosis in these ischemic syndromes using caspase inhibition. Complementary pharmacologic (peptide pseudosubstrates) and transgenic (overexpression of a dominant caspase inhibitor) approaches will be employed. 3. To determine the contribution of apoptosis to changes in myocardial structure and function during and after infarction and ischemia-reperfusion injury. Using caspase inhibition, the contribution of myocyte apoptosis to infarct size, ventricular remodeling, and contractile dysfunction will be determined. These studies will increase our understanding of the mechanism of cardiac myocyte apoptosis and its role in the pathogenesis of ischemic heart disease.